Cardiovascular disease is the most frequent cause of death in the United States. Of the many cardiovascular diseases, myocardial infarction (MI) and stroke are identified as the most prominent in sudden onset syndromes, both requiring the need of immediate and urgent medical care. Critical to the rapid intervention of such cardiac events are point of care assays able to accurately determine the onset of a true event so that hospitalization and treatment are administered in an effective and economical manner. Regarding myocardial infarction (MI), such testing generally focuses on an established cardiac biomarker panel consisting of creatine kinase MB (CKMB), cardiac troponin (cTn) and myoglobin (MYO), as monitored from plasma/serum. Notably, these markers follow independent temporal curves, with each reaching its greatest clinical/diagnostic usefulness in time frames ranging from 2-to-24 hours post onset of chest pain. The generally reported predictive values of these biomarkers are CK-MB; sensitivity=99%, specificity=89%, Positive Predictive Value (PPV)=64%, Negative Predictive Value (NPV)=99.8%; Troponin I; sensitivity=33.3%, specificity=99.4%, PPV=78.6%, NPV=96.6%; MYO; sensitivity=84.6%, specificity=71.1%, PPV=20.4%, NPV=98.3%.
Of particular interest is the earliest responding marker, myoglobin, which may be used to establish negative predictive assessments—i.e., for ruling out MI—within ˜3-hours of suspected onset. Used as a single marker with a reference level of >55-200 ng/mL (dependent upon the sex of the patient and the commercial assay in use), myoglobin is used to assess MI patients with reported PPV=20.4% and NPV=98.3%, (with clinical sensitivity and specificity of 84.6% and 71.1%, respectively) within 3 hours of admission into an emergency department. To improve the accuracy of prediction in MI candidates, myoglobin is often used in combination with either or both of the other established markers. Predictive values of PPV=19.0 and NPV=99.5 have been reported in studies using myoglobin in combination with one or both of CKMB and cTn in the same 3-hour time frame. These studies imply that the preferred means of MI determination is through multiple-marker assays employed at the earliest possible time after suspecting onset.
However, it is accurate to state that the predictive metrics of the conventional MI markers are far from perfect, regardless of whether used individually or in various combinations. Thus, there clearly exists a need for additional and improved high sensitivity and specificity tests able to rapidly differentiate between MI and complicating symptoms. Other than patient-in/patient-out determinations, these same tests are called upon as follow-through monitors able to gauge the effectiveness of treatment and detect the onset of secondary MI events. For these foregoing reasons, there is a pressing need to discover, identify and validate new biomarkers denoting the occurrence of MI as well as a need to improve the clinical sensitivity/specificity of existing cardiovascular biomarkers.
Described in this invention are new biomarkers found to be indicative of MI and the subsequent design of assays, including multiple-marker mass spectrometric immunoassays (MSIA), tailored for the determination of MI. Specifically, this invention describes newly discovered cardiovascular disease biomarkers; serum amyloid A and its variants (collectively referred to as SAA), and transthyretin and its variants (collectively referred to as TTR). As used herein, protein variants include but are not limited to, proteins having posttranslational modifications or point mutations, proteolytic or chemical fragments of the protein, truncated forms of the protein, the protein in multimeric complexes, or any of the aforementioned in combination. Of particular importance are the SAA variant; intact wild-type SAA1α (SAA1α) and the TTR variant sulfo-TTR (SS-TTR). Provisions are made to use these newly discovered cardiovascular biomarkers individually or in various combinations with themselves and/or other proteins for the accurate determination of MI. Also provided for is a multiplexed assay using variants of TTR, variants of SAA and variants of an existing MI biomarker, myoglobin (collectively referred to as MYO), to accurately determine MI.
Accordingly, this invention makes provisions for the concept of finding, identifying and using new biomarkers that are able to improve the clinical and diagnostic monitoring of cardiovascular diseases. Moreover, this invention makes provisions for designing and implementing assays able to analyze one or more markers found to be associated with MI. Finally, the present invention makes provisions for an analytical platform able to perform these assays in a multiplexed manner, and means of correlating the data (that include threshold detection and computer-assisted correlations) for improved clinical and diagnostic utility.